Histopathology
of
cadmium
toxicitya.
Treated
male
mice
(5-4
per
strain
per
study
group)
are
injected
with
15
µmol
CdCl2/kg
(20
µCi
109Cd/kg)
i.v.,
dissolved
in
physiological
saline
(0.9%
NaCl)
in
a
volume
of
10
mL/kg
BW
bolus,
via
the
tail
dorsal
vein,
while
the
control
group
is
given
saline
vehicle.
b.
After
24
hrs
following
CD
administration,
the
mice
are
deeply
anesthetized
with
60
mg/kg
BW
dose
of
pentobarbital.
c.
Under
adequate
anesthesia,
the
thoracic
cavity
is
opened
for
total
body
perfusion
via
the
heart,
initially
with
normal
saline
to
flush
the
blood,
and
then
with
freshly
made
0.4%
paraformaldehyde
for
pre-fixation.d.
The
perfusion
pre-fixed
brains,
trigeminal
ganglias,
liver,
and
testes
are
dissected,
weighed,
and
Cd
content
determined
using
gamma
scintillation
spectrometry.e.
Following
Cd
content
ditermination,
the
organs
are
placed
in
fresh
0.4%
paraformaldehyde
or
10%
neutral
formalin
for
24
h.
f.
Fully
fixed
liver,
ganglia,
and
testes
are
then
processed
via
standard
histopathological
techniques,
sectioned
at
5
µm
thickness,
and
stained
with
H&E.
g.
Three
sections
from
the
each
entire
trigeminal
ganglion,
liver,
and
testes
(9
sections
from
each
mouse)
are
microscopically
evaluated
and
scored
for
cadmium
toxicity
(see
Figures
1-4
below).
h.
Liver
sections
are
evaluated
for
necrosis,
apoptosis,
and
other
lesions;
hepatocytes
(1500–2500
per
mouse)
in
12–20
randomly
selected
fields
are
counted
under
400X
magnification,
aided
by
a
grid
of
100
squares
(see
Figures
1-2
below).
i.
Apoptotic
index
(AI)
and
mitotic
index
(MI)
are
determined
by
dividing
the
total
number
of
hepatocytes
showing
apoptosis
(apoptotic
cells
and
bodies)
or
mitosis
by
the
total
number
of
cells
in
the
fields
examined.
Clusters
of
two
or
three
apoptotic
bodies
are
regarded
as
single
apoptotic
events
(Habeebu
et
al,
1998).
j
.
Typing
of
apoptotic
cells
and
bodies
(intracellular
or
extracellular,
chromatin-containing
or
non-chromatin-containing)
is
done
at
1000X
magnification
or
100X
objective
(with
oil
immersion).
k.
Necrosis
is
analyzed
semiquantitatively
with
five
scores
for
severity:
0=
none;
1=
necrosis
of
1–5%
of
hepatocytes;
2=necrosis
of
6–25%
of
hepatocytes;
3=necrosis
of
26–50%
of
hepatocytes;
and
4=necrosis
of
>50%
of
hepatocytes.

Figure
1.
A
schematic
example
of
a
partially
necrotic
liver-partly
overlayed
with
a
scoring
grid.